1. Field of the invention:
This invention relates to a method for measuring the amount of polyamines in a biological sample, and in particular, to a method for measuring the amount of polyamines in an urine sample, in which the polyamines can be measured accurately without being affected by interfering substances that may be present in the biological sample.
2. Description of the prior art:
The polyamines (including diamines) that are found in fluids from the body such as urine are known to be closely correlated to the onset of cancer and to the state of its progress. Eleven polyamines have been identified, including cadaverine, putrescine, spermidine, and spermine. Also, acylated forms of the said polyamines, i.e., acylpolyamines, have also been found. The measurement of total polyamines (below, the word "polyamines" is meant to include acylpolyamines) in fluids from the body and also the measurement of their individual levels have much clinical significance, because the values have been suggested to give information useful for the diagnosis of cancer. For that reason, a method is needed in which the amounts of polyamines can be measured with a high degree of accuracy.
In the conventional methods for measuring the amount of polyamines, polyamines in biological samples are isolated by the use of liquid chromatography or electrophoresis, and then measurement of the polyamines is done by a fluorescence method or by a colorimetric method with ninhydrin. However, these methods require the troublesome steps of pretreatment of the biological samples, and much time is needed for the measurement. For that reason, these methods cannot be used for carrying out the measurement of a large number of biological samples at once. In addition, these methods require special techniques, equipment, and facilities. Therefore, these methods cannot be used as, for example, routine procedures in clinical laboratories.
Japanese Laid-Open Patent Publication 56-36918 discloses a relatively simple method for measuring the amount of polyamines. In the method, polyamines are reacted with amine oxidase to produce hydrogen peroxide, and the hydrogen peroxide is reacted with 4-aminoantipyrin, phenol and peroxidase resulting in a pigment. The pigment is measured by colorimetric analysis. In this method, there is no need to isolate the polyamines from the biological samples in pure form by the use of liquid chromatography or electrophoresis. However, in this method, error arises because of the reducing substances present in the biological sample, so it is necessary to remove the reducing substances from the biological samples beforehand. For example, it is necessary to remove such reducing substances from the biological sample by passage of the biological sample through a column filled with a cation-exchange resin. Because of this pretreatment step, the polyamines in the biological sample cannot be measured in a single step by the use of an automated analyzer.